The long term objective is to identify and characterize the various components of taste transduction pathways to understand how they function in the taste cell. Of particular importance are taste cell guanine nucleotide binding regulatory proteins (G proteins) and the seven transmembrane helix receptors that couple to them. G proteins regulate effector enzymes such as phosphodiesterase and phospholipase to effect taste cell changes in intracellular second messengers (e.g., cAMP, cGMP, IP3, Ca2+). Gustducin is a taste specific G protein closely related to the transducins (the photoreceptor G proteins). Recently, it has been shown that rod transducin is also expressed in taste cells. These findings suggest that taste transduction may have some similarities to phototransduction. The specific goals of the proposal are the following: 1) to characterize gustducin's role in taste transduction using biochemical methods; 2) to purify, physically characterize and molecularly clone a recently identified taste specific phosphodiesterase (PDE); 3) to characterize the gustducin-PDE interaction using peptides and mutated gustducin and transducin proteins; 4) to characterize gustducin's role in taste transduction using behavioral and electrophysiological analysis of transgenic mice. Preliminary data indicate significant differences between gustducin knock out and wild type mice in their behavioral and electrophysiological responses to both sweet and bitter compounds; 5) the murine gustducin promoter will be used to express gustducin and transducin as transgenes in the taste cells of gustducin knock out mice. If transducin rescues knock out mice, it will indicate that gustducin and transducin are interchangeable in taste transduction; 6) transgenic mice will be made that express mutant gustducin deficient in its interaction with PDE. These transgenic animals will be tested behaviorally and electrophysiologically to determine if mutant gustducin also leads to altered bitter and sweet responses. Diverse methods will be required to address these goals: biochemical, transgenic, behavioral and electrophysiological techniques will be used. the results of these studies will provide significant new insights into the molecular mechanisms underlying taste transduction. Gustatory and metabolic disorders such as malgeusia, dysgeusia and cachexia frequently occur in conjunction with several types of cancer. The knowledge gained from this proposal may further our understanding of the molecular bases of taste disorders and eventually lead to effective intervention.